The present invention relates to the treatment and prevention of diseases caused by protozoan parasites in man and in animals. In particular, the invention relates to novel compositions and methods for parenteral treatment and prevention of protozoal diseases, e.g., Piroplasmosis, Babesosis, Toxoplasmosis Neospora caninum, Crytosporidiosis and Equine Protozoal Myeloencephalitis.
Protozoan parasites (also known as apicomplexan parasites) cause a variety of clinical disease manifestations in both man and animals. For example, hemoprotozoan parasites of the Babesia genus, namely Babesia caballi and Babesia equi, are responsible for the economically devastating disease, equine piroplasmosis. Equine piroplasmosis is widely distributed worldwide although it is most prevalent in the tropics, sub-tropics and temperate regions (see, Robinson, Edward N., xe2x80x9cCurrent Therapy In Equine Medicinexe2x80x9d, Vol.2, pp.299-300, (1987) (ISBN: 0-7216-14914)).
The principal mode of transmission of the protozoan is via a tick vector, e.g., Dermacentor nitens. Clinical manifestation of acute infection is characterized by depression, fever, anorexia, icteric mucous membranes, ecchymotic hemorrhages and edema of the extremities and ventral abdomen. Death can occur within 24-48 hours and mortality rates in outbreaks may be high (see, Robinson, Edward, N., xe2x80x9cCurrent Therapy in Equine Medicinexe2x80x9d, Vol. 3, pp. 499-500 (1992) (ISBN: 0-7216-3475-3)).
Horses which test positive for piroplasmosis (complement fixation or indirect flourescent antibody tests) are rarely allowed to enter the United States without first undergoing treatment for the disease or under rigid guidelines (see, Brooks, L., xe2x80x9cPiroplasmosis: The Olympic Questionxe2x80x9d, The Horse, pp. 43-48 (July 1996)). Currently recommended treatment regimens include the use of imidocarb dipropionate (Burroughs Welcome Co.) and often the treatment produces adverse side effects which include salivation, restlessness, colic and gastrointestinal tract hypermotility (see, Kobluk, Calvin N. et al., xe2x80x9cThe Horse Diseases and Clinical Managementxe2x80x9d, Vol. 2, pp. 1084-1885 (1995) (ISBN: Vol.2 0-7216-5984-5)). Moreover, treatment with imidocarb dipropionate has met with marginal success, especially when the etiologic agent is Babesia equi (50% to 60%) (see, Reed, Stephen M. et al., xe2x80x9cEquine Internal Medicinexe2x80x9d, pp. 570-571 (1998) (ISBN: 0-7216-3524-5)).
Thus, there is a need in the art for an effective anti-protozoal agent and a method of treatment and prevention of Piroplasmosis which does not produce the adverse side effects seen with current treatment regimens.
Other examples of blood borne protozoal infection caused by Babesia spp. include: babesiosis of cattle, e.g., Babesia bigemina and Babesia bovis; caprine and ovine babesiosis, e.g., Babesia ovis; and canine babesiosis, e.g., Babesia canis and Babesia gibsoni (see, Smith, Bradford P., xe2x80x9cLarge Animal Internal Medicinexe2x80x9d pp.1088-1092 (1990) (ISBN: 0-8016-5062-3)). See also, Bonagura, John D. xe2x80x9cKirk""s Current Veterinary Therapy XII Small Animal Practicexe2x80x9d, Vol. 12, pp. 315-319, (1995) (ISBN: 0-7216-5188-7)). Likewise, there is a need in the art for a safe, effective and economical treatment for such infections.
In humans, for example, protozoan infections can cause severe disease manifestations. A common sequella in patients suffering from acquired immune defficiency syndrone (AIDS) is Cryptosporidium parvum infection (Cryptosporidiosis) which produces severe chronic and often fatal diarrhea. The parasite is found worldwide and lives in cattle and domestic animals and is excreted in feces. It can be transmitted to humans directly from animals or through contact with feces, contaminated water or food (see, e.g. xe2x80x9cNational Institute of Allergy and Infectious Diseasesxe2x80x94AIDS-Related Cryptosporidiosisxe2x80x9d, www press release (March, 1991)).
Many attempts have been made to find a threapeutically effective treatment for this disease. One family of drugs currently used in the veterinary profession for the treatment of coccidosis, the triazine-based anticoccidial agents (e.g., triazinediones and triazinetriones) especially diclazuril and letrazuril, have been tried experimentally in the treatment of crytosporidiosis in man (see, National Library of Medicine, AIDSDRUGS Database, DRG-0079 (Jan. 22, 1998); and (National Library of Medicine, AIDSTRIALS Database, FDA-038B (Apr. 25, 1990)). These compounds are formulated for oral administration and have met with limited success due to poor absorption. The best response to such drugs has been seen in persons with the highest blood levels post adminsitration (see, AIDS Treatment News, No. 111 (Sep. 21, 1991)).
To date, however, there is still no efficacious therapy for Cryptosporidial infections in man (see, Health Canada, Laboratory Centre For Disease Control: Material Safety Data Sheetxe2x80x9448, Cryptosporidium parvum, Oct. 11, 1997 @ (www.hc-sc.gc.calhpb/lcdc/biosafty/msds/msds/48e.html (Jul. 24, 1999)). Accordingly, there still exists an urgent need in the art for a safe and effective pareneral formulation for the treatment and prevention of potozoal infections such as cryptosporidiosis or babesiosis.
Equine protozoal myeloencephalitis (EPM), a central nervous system disease which affects equine species, is also primarily caused by a protozoan parasite, Sarcocystis neuroma also known as Sarcocystis falcatula. The horse is not a normal host for this protozoan (the horse is not part of the normal life cycle) and is considered to be a dead end host. The definitive host is thought to be the opossum. Equids are infected with the S. neuroma organisms via ingestion of food or water contaminated with feces of an infected carnivore such as the opossum (see, Robinson, Edward N., Current Therapy in Equine Medicine: Fenger, Clara A., xe2x80x9cEquine Protozoal Myeloencephalitisxe2x80x9d, Vol. 4, pp. 329-333 (1997) (ISBN: 0-7216-2633-5)).
Recently, other protozoan parasites have been implicated as also playing an etiologic role in the pathogenesis of EPM, e.g., Neospora caninum and Toxoplasma species. Accordingly, there still exists a need in the art for an effective treatment of EPM which demonstrates broad spectrum efficacy against all protozoan parasites in the horse inclusive of the aforementioned organisms.
The clinical signs of EPM can vary from case to case. Generally, horses present neurological signs which are asymmetrical, and actual symptoms will vary depending upon the severity and location of lesions produced by the parasites in the brain, brain stem or spinal cord. Ataxia, incoordination and general weakness are usually present and can be accompanied by muscle atrophy (usually most notable in the rear limbs). There can be paralysis of the muscles of the eyes and face, drooping ears, difficulty swallowing (dysphagia) head tilt, altered gait, or even seizures and collapse. Recent reports of numbers of EPM cases indicate that the disease is far more widespread and serious than originally thought.
There is currently no vaccine available for prevention of this disease. Previously preferred treatment was aimed at control of the parasitic infection via the use of sulphonamides and pyrimethamine (see, U.S. Pat. No.: 5,747,476). However, these measures have been met with limited success. More recently, and due to the urgent need for a safe and effective treatment for this devastating disease, new methods of therapy and new agents have been explored, e.g., through the emergency FDA importation of anticoccidial agents such as diclazuril and toltrazuril (see, FDACVM publication: xe2x80x9cInstructions for Personal Use Importation of Diclazurilxe2x80x9d (Dec. 16, 1997); and FDACVM publication: xe2x80x9cThe Importation of Toltrazuril for Personal Usexe2x80x9d (1997) which are available from the American Association of Equine Practitioners, Lexington, Ky. (AAEP)). (See also, U.S. Pat. No.:5,883,095).
It should be noted that the current emergency importation and treatment of horses with diclazuril and toltrazuril is speculative. Oral formulations adapted for use in the horse of one or more of these drugs are currently in clinical trials for FDA approval. And, while some horses do dramatically improve, many only see moderate improvement (improve 1-2 grades on the clinical evaluation scale (ranging in grades of 1-5) used by veterinarians to classify the severity of clinical signs). The test dose for a presently preferred form of toltrazuril, the metabolite toltrazuril sulfone (Bayer) is currently 5 mg/kg up to 10 mg/kg per day for the FDA trials.
The cost for importation and use of the above anticoccidials ranges anywhere from about $800-$1,200.00 per horse and results obtained from the treatment can be disheartening. Few horses experience complete recovery from any known conventional therapy, including the triazine-based anticoccidial therapies which are mentioned above. Moreover, the relatively high levels of drug recommended in the therapeutic regimen can produce unwanted side effects. This is especially true for treatment regimens which utilize sulfonamides and pyrimethamine which inhibit folic acid production (see, Fenger, Clara A., xe2x80x9cUpdate on the Diagnosis of Equine Protozoal Myeloencephalitis (EPM)xe2x80x9d Proc. 13th ACVIM Forum, pp.597-600 (1995); and Bertone, Joseph J., xe2x80x9cUpdate On Equine Protozoal Myeloencephalitisxe2x80x9d, FDA Veterinarian, Vol. XI, No. III (May/June 1996)).
Therefore, there exists a need in the art to provide a safe and more effective method of parenteral treatment of EPM which is also less costly.
The present invention satisfies the need in the art by providing a composition containing at least one anti-protozoal drug especially adapted for parenteral administration, e.g., intranasal, intramuscular, subcutaneous, transdermal or intraveneous administration, for the treatment and prevention of protozoan infections in man an in animals. In one embodiment, the anti-protozoal drug is a triazine-based anticoccidial agent, e.g., a triazinedione or triazinetrione such as diclazuril or toltrazuril. In a presently preferred embodiment, the triazine-based anticoccidial agent is sulfonotoltrazuril.
The compositions of the invention can further comprise a suitable solvent for the anti-protozoal drug in a formulation specially adapted for a particular route of parenteral administration. The choice of solvent and concentration of active dissolved therein will of course vary depending upon the choice of drug, the desired route of parenteral administration, the species and host being treated, and the desired duration of action of the administration, e.g., sustained release vs. loading dose.
Parenteral administration of the compositions of the invention reduces the dosage amount of drug by about one fold to as much as one hundred fold but especially about five fold compared with oral dosing. The compositions provided by the invention eliminate the variability in plasma concentrations of the drug due to animal to animal differences in oral bioavailability, allow use of a loading dose and therefore immediate attainment of effective plasma concentrations of the drug, allow rapid attainment of high plasma concentration of the drug to drive the drug into extravascular compartments such as the cerbrospinal fluid in the CNS, provide better and more immediate control of the plasma concentration of the drug and reduce the potential for side effects related to the currently existing oral formulations. As a result, the cost of treatment is greatly reduced along with the potential for adverse side effects as seen from higher doses currently recommended and needed for oral administration of these anti-protozoal drugs.
In particular, the present invention provides novel compositions and methods for the treatment and/or prevention of any anti-protozoal or apicomplexan parasite in man and animals, e.g. equine piroplasmosis, equine protozoal myeloencephalitis and Cryptosporidiosis. In one embodiment, the invention provides a composition comprised of a triazine-based anticoccidial drug and a suitable solvent useful for the treatment of anti-protozoal infections. A preferred embodiment of the invention comprises a composition comprised of diclazuril in solution with DMSO, DMA or mixtures thereof which is formulated for parenteral administration for the treatment of anti-protozoal infection in man or in animals as provided by the methods of the invention.
Another preferred embodiment of the invention comprises a composition comprised of toltrazuril, toltrazuril sulfone, sulfonotoltrazuril or mixtures thereof in solution with DMSO, DMA or mixtures thereof which is formulated for parenteral administration for the treatment of anti-protozoal infection in man or in animals as provided by the methods of the invention.
Yet another preferred embodiment of the invention comprises a composition formulated for parenteral or oral administration for treatment of anti-protozoal infections in man and in animals comprised of a soluble salt, e.g., a sodium salt, of a triazine-based anticoccidial agent, e.g., clazuril, diclazuril, letrazuril, toltrazuril, toltrazuril sulfone, or sulfonotoltrazuril and the like.
Also provided by the invention is a method for preparation of water-soluble forms of triazine-based anticoccidial agents, e.g., clazuril, diclazuril, letrazuril, toltrazuril, toltrazuril sulfone, or sulfonotoltrazuril for use in the methods of treatment of anti-protozoal infections provided by the invention.
The present invention provides a composition and method for the parenteral treatment of protozoan (apicoplexan) infections in man and in animals. The protozoan parasite can be any protozoan known to infect man or animals, including, but not limited to, e.g., Babesia spp. Sarcocystis spp. Neosporum spp. Crytosporidium spp. Toxoplasma spp. and the like.
It is contemplated that the compositions of the invention can be formulated for any parenteral administration. It is specifically contemplated that intravenous, intramuscular, transdermal, intranasal and subcutaneous routes of administration can be utilized for administration of the compositions of the invention. Specific formulations of the compositions of the invention can include powders, gels, ointments, creams, solutions, suspensions, sustained release preparations, patches and the like.
In one embodiment, the invention provides a composition specially adapted for intravenous, intramuscular, subcutaneous, or intranasal administration which is useful for the treatment of a protozoal infection in man or in animals, e.g., for treatment of equine piroplasmosis, equine protozoal mycloencephalitis or human cyrptosporidiosis infection, wherein the composition comprises at least one chemical agent which has anti-protozoal activity. The compositions provided herein can include any anti-protozoal agent, but especially anti-coccidial agents such as any of the class of triazine-based anti-coccidial agents (i.e., agents which contain a triazine ring, e.g., the 1, 2, 4 triazine ring or the 1, 3, 5 triazine ring configurations (see, e.g. xe2x80x9cA 3D-QSAR Study of Anticoccidial Triazines Using Molecular Shape Analysisxe2x80x9d, J. Chem. Inf. Comput Sci., Vol. 35, 771-778 (1995); U.S. Pat. No. 4,837,216; and U.S. Pat. No. 4,952,570, the contents of which are incorporated herein by reference).
Specific examples of such agents include, but are not limited to, clazuril, diclazuril, toltrazuril, toltrazuril sulfone, or sulfonotoltrazuril. For example, the chemical structures of several triazine-based compounds useful in the compositions and methods set forth herein are shown below: 
It can be appreciated that other anti-protozoal agents including their derivatives, analogs, isomers, salts, and natural metabolites of these agents can also be utilized in the compositions for parenteral treatment and prevention of any protozoal infection in man or in animals. In a presently preferred embodiment, the triazine-based anticoccidial agent is sulfonotoltrazuril, a toltrazuril metabolite.
Sufonotoltrazuril, the toltrazuril metabolite, contains the thio group of toltrazuril that has been oxidized to a sulfono group thereby differing toltrazuril from its dioxo sulfonotoltrazuril derivative. 
The chemical name used in the Chemical Abstract Database for one form of the sulfonotoltrazuril contemplated by the invention is: 1-methyl-3-[3-methyl-4-(4-trifluoromethanesulfonyl-phenoxy)-phenyl]-[1,3,5]triazinane-2,4,6-trione.
The CAS catalog number of this compound is CAS No. 69004-04-2. The Beilstein Registry Number is 870959. The Molecular Formula is C18H14F3N3O6S and the Molecular Mass is 457.38 g/mol.
In particular, the invention provides a composition which is useful for parenteral treatment and/or prevention of EPM which utilizes between about xc2xcth to about {fraction (1/100)}th of the amount of an anti-protozoal drug which is necessary for oral administration in the treatment of EPM. In one embodiment of the invention, the compositions are adapted for intranasal administration and comprise about {fraction (1/10)}th of the currently recommended dosage. In another embodiment, the compositions are adapted for other parenteral administration (e.g., intravenous, subcutaneous and intramuscular) and comprise between about xc2xcth to about {fraction (1/100)}th of the currently recommended oral dosage, but especially about ⅓rd to about {fraction (1/10)}th of the currently recommended oral dosage for EPM treatment. In a preferred embodiment, the intranasal and parenteral compositions of the invention set forth above are formulated for sustained release as set forth in greater detail below.
For example, the currently recommended dosage for oral diclazuril (CLINACOX(copyright), Pharmacea Upjohn, Canada) in the horse for an EPM treatment regimen is about 2.5 grams of diclazuril per 1000 pound horse per day (5.5 mg/kg) administered once daily for 28 days. This amounts to about 70 grams of diclazuril per horse per treatment regimen.
By contrast, the presently preferred dosage range for the parenteral compositions of the present invention for treatment of EPM which are comprised of similar triazine-based agents, e.g., diclazuril or toltrazuril and the like, is from between about 0.1 mg/kg to about 10 mg/kg. However, the skilled artisan can appreciate that this range can vary from between about 0.01 mg/kg to about 20 mg/kg depending upon the specific formulation, route of administration, the desired effect (loading dose vs. sustained release) and the duration of the treatment regimen.
A presently preferred embodiment of the invention comprises a composition adapted for parenteral administration wherein the anti-protozoal agent is selected from, but not limited to the group consisting of clazuril, diclazuril, letrazuril, toltrazuril, toltrazuril sulfone, and sulfonotoltrazuril or a sodium salt thereof and a suitable solvent. The solvent can be any suitable solvent for use in animals and man and will, of course vary depending upon the choice of anti-protozoal agent and the route of administration. Presently preferred solvents include, but are not limited to DMSO, DMA, ethanol, water and the like as set forth more fully below.
The preferred compositions can be utilized in methods of treatment of anti-protozoal infections in man and in animals. For example, a presently preferred treatment regimen for treatment of EPM comprises administering to a 1000 lb horse a composition for intravenous administration comprised of between about 50 mg to about 1,500 mg , but especially between about 250 mg and about 1000 mg and most preferably about 500 mg (about 1.1 mg/kg) of diclazuril, toltrazuril, toltrazuril sulfone or sulfonotoltrazuril dissolved in a suitable volume of DMSO, DMA or the like. Suitable amounts of solvent will vary from between about 2 ml to about 30 ml per unit dose depending upon the choice of the anti-protozoal agent and the choice of solvent. According to the methods set forth herein, the composition can given once per day (SID) for a duration of between about 10 and about 35 days but especially between about 20 and about 30 days and most preferably about 28 days. Alternatively, a loading dose of the composition can be given to achieve rapid critical plasma concentrations on day one of the regimen followed by a maintenance dose (see, Example 1 below) for a shorter duration of therapy, e.g., between about 15 and about 25 days.
In yet another embodiment, the compositions can be specially formulated for parenteral use, e.g., intramuscular or subcutaneous, sustained release, e.g., a microsphere or methylcellulose preparation such that a single sustained release administration or a weekly administration of sustained a sustained release fonrulation of the anti-protozoal agent is possible. Alternatively, a single intraveneous loading dose followed by a sustained release intramuscular or subcutaneous dose for maintaining sustained critical blood levels is contemplated. In EPM, for example, the critical plasma concentration can range from about 5 xcexcg/ml of the antiprotozoal agent to about 12 xcexcg/ml, but especially about 8 xcexcg/ml.